A Distinct Mechanism to Achieve Efficient Signal Recognition Particle (SRP)-SRP Receptor Interaction by the Chloroplast SRP Pathway

Cotranslational protein targeting by the signal recognition particle (SRP) requires the SRP RNA, which accelerates the interaction between the SRP and SRP receptor 200-fold. This otherwise universally conserved SRP RNA is missing in the chloroplast SRP (cpSRP) pathway. Instead, the cpSRP and cpSRP receptor (cpFtsY) by themselves can interact 200-fold faster than their bacterial homologues. Here, cross-complementation analyses revealed the molecular origin underlying their efficient interaction. We found that cpFtsY is 5- to 10-fold more efficient than Escherichia coli FtsY at interacting with the GTPase domain of SRP from both chloroplast and bacteria, suggesting that cpFtsY is preorganized into a conformation more conducive to complex formation. Furthermore, the cargo-binding M-domain of cpSRP provides an additional 100-fold acceleration for the interaction between the chloroplast GTPases, functionally mimicking the effect of the SRP RNA in the cotranslational targeting pathway. The stimulatory effect of the SRP RNA or the M-domain of cpSRP is specific to the homologous SRP receptor in each pathway. These results strongly suggest that the M-domain of SRP actively communicates with the SRP and SR GTPases and that the cytosolic and chloroplast SRP pathways have evolved distinct molecular mechanisms (RNA vs. protein) to mediate this communication

Concerted Complex Assembly and GTPase Activation in the Chloroplast Signal Recognition Particle

The universally conserved signal recognition particle (SRP) and SRP receptor (SR) mediate the cotranslational targeting of proteins to cellular membranes. In contrast, a unique chloroplast SRP in green plants is primarily dedicated to the post-translational targeting of light harvesting chlorophyll a/b binding (LHC) proteins. In both pathways, dimerization and activation between the SRP and SR GTPases mediate the delivery of cargo; whether and how the GTPase cycle in each system adapts to its distinct substrate proteins were unclear. Here, we show that interactions at the active site essential for GTPase activation in the chloroplast SRP and SR play key roles in the assembly of the GTPase complex. In contrast to their cytosolic homologues, GTPase activation in the chloroplast SRP–SR complex contributes marginally to the targeting of LHC proteins. These results demonstrate that complex assembly and GTPase activation are highly coupled in the chloroplast SRP and SR and suggest that the chloroplast GTPases may forego the GTPase activation step as a key regulatory point. These features may reflect adaptations of the chloroplast SRP to the delivery of their unique substrate protein

Transient tether between the SRP RNA and SRP receptor ensures efficient cargo delivery during cotranslational protein targeting

Kinetic control of macromolecular interactions plays key roles in biological regulation. An example of such control occurs in cotranslational protein targeting by the signal recognition particle (SRP), during which the SRP RNA and the cargo both accelerate complex assembly between the SRP and SRP receptor FtsY 10^2-fold. The molecular mechanism underlying these rate accelerations was unclear. Here we show that a highly conserved basic residue, Lys399, on the lateral surface of FtsY provides a novel RNA tetraloop receptor to mediate the SRP RNA- and cargo-induced acceleration of SRP–FtsY complex assembly. We propose that the SRP RNA, by using its tetraloop to interact with FtsY–Lys399, provides a transient tether to stabilize the early stage and transition state of complex formation; this accelerates the assembly of a stable SRP–FtsY complex and allows the loading of cargo to be efficiently coupled to its membrane delivery. The use of a transient tether to increase the lifetime of collisional intermediates and reduce the dimension of diffusional search represents a novel and effective mechanism to accelerate macromolecular interactions

Cryo-EM structure of the E. coli translating ribosome in complex with SRP and its receptor

We report the 'early' conformation of the Escherichia coli signal recognition particle (SRP) and its receptor FtsY bound to the translating ribosome, as determined by cryo-EM. FtsY binds to the tetraloop of the SRP RNA, whereas the NG domains of the SRP protein and FtsY interact weakly in this conformation. Our results suggest that optimal positioning of the SRP RNA tetraloop and the Ffh NG domain leads to FtsY recruitment

The Intervening Role of Urgency on the Association Between Childhood Maltreatment, PTSD, and Substance-Related Problems

A range of risk factors lead to opioid use and substance related problems (SRP) including childhood maltreatment, elevated impulsivity, and psychopathology. These constructs are highly interrelated such that childhood maltreatment is associated with elevated impulsivity and trauma-related psychopathology such as posttraumatic stress disorder (PTSD), and impulsivity—particularly urgency—and PTSD are related. Prior work has examined the association between these constructs and SRP largely independently and it is unclear how these multi-faceted constructs (i.e., maltreatment types and positive and negative urgency) are associated with one another and SRP. The current study used structural equation modeling (SEM) to examine the relations among childhood maltreatment types, trait urgency, PTSD symptoms, and SRP in a sample of individuals with a history of opioid use. An initial model that included paths from each type of childhood maltreatment, positive and negative urgency, PTSD and SRP did not fit the data well. A pruned model with excellent fit was identified that suggested emotional abuse, positive urgency, and negative urgency were directly related to PTSD symptoms and only PTSD symptoms were directly related to SRP. Furthermore, significant indirect effects suggested that emotional abuse and negative urgency were related to SRP via PTSD symptom severity. These results suggest that PTSD plays an important role in the severity of SRP among those with significant maltreatment histories and negative urgency

Signal recognition particle binds to translating ribosomes before emergence of a signal anchor sequence.

The bacterial signal recognition particle (SRP) is part of the machinery that targets ribosomes synthesizing membrane proteins to membrane-embedded translocons co-translationally. Recognition of nascent membrane proteins occurs by virtue of a hydrophobic signal-anchor sequence (SAS) contained in the nascent chain, usually at the N terminus. Here we use fluorescence-based stopped-flow to monitor SRP-ribosome interactions with actively translating ribosomes while an SRP substrate is synthesized and emerges from the peptide exit tunnel. The kinetic analysis reveals that, at cellular concentrations of ribosomes and SRP, SRP rapidly binds to translating ribosomes prior to the emergence of an SAS and forms an initial complex that rapidly rearranges to a more stable engaged complex. When the growing peptide reaches a length of ∼50 amino acids and the SAS is partially exposed, SRP undergoes another conformational change which further stabilizes the complex and initiates targeting of the translating ribosome to the translocon. These results provide a reconciled view on the timing of high-affinity targeting complex formation, while emphasizing the existence of preceding SRP recruitment steps under conditions of ongoing translation

A nonlinear high temperature fracture mechanics basis for strainrange partitioning

A direct link was established between Strainrange Partitioning (SRP) and high temperature fracture mechanics by deriving the general SRP inelastic strain range versus cyclic life relationships from high temperature, nonlinear, fracture mechanics considerations. The derived SRP life relationships are in reasonable agreement based on the experience of the SRP behavior of many high temperature alloys. In addition, fracture mechanics has served as a basis for derivation of the Ductility-Normalized SRP life equations, as well as for examination of SRP relations that are applicable to thermal fatigue life prediction. Areas of additional links between nonlinear fracture mechanics and SRP were identified for future exploration. These include effects of multiaxiality as well as low strain, nominally elastic, long life creep fatigue interaction